Jun 09, 2022
July 28, 2022
RNA vaccines are hugely versatile. They can potentially inoculate against any pathogen for which a protein target is known, for example, influenza, chlamydia, HIV-1 (human immunodeficiency virus 1), Ebola and RSV (respiratory syncytial virus). Self-amplifying RNA (saRNA) maintains the advantages of mRNA vaccines, such as rapid development, modular design, and cell-free synthesis, but only requires a lower dose of RNA due to the self-replicative properties while maintaining its efficacy and protection against disease. This reduces the burden of manufacturing, allowing facilities to make 100x more doses than with mRNA, lowering cost and time. As a result, distributed /decentralized manufacturing and quick mass immunizations are possible in the event of a pandemic.
Challenges for RNA-based drug or vaccine
The two challenges to the successful development of RNA vaccines are:
1. RNA is a large, negatively charged molecule and requires a carrier vehicle to get into the cells.
2. RNA is extremely delicate and is likely to break down quickly during process development, manufacturing and or during formulation development.
Unlike many small molecule and biologic drugs, nucleic acid therapeutics, including saRNA, are ineffective as unformulated molecules and thus require delivery technology to avoid degradation and protect the cargo to ensure efficacy when it arrives at the target cells. Hence, systematically screening various delivery vehicles and selecting appropriate formulations are critical to developing next-generation mRNA vaccines.
The Solution - Lipid nanoparticle (LNP) technology meets the delivery need for saRNA vaccine
The philosophy behind safe and efficient delivery systems is to condense the saRNA in a cationic carrier that can be taken into the cell without being degraded. So far, delivery platforms for saRNA have included lipid nanoparticles (LNP), polyplexes and cationic nanoemulsions (Figure 2). Among all the delivery platforms, LNP is the most clinically advanced, with the recent FDA (Food and Drug Administration) approval of COVID-19 based-modified mRNA vaccines. At present, the saRNA vaccine has been used in vaccine development against many infectious diseases like malaria and COVID-19 and several saRNA LNP vaccine clinical trials are currently underway.
The Lipid nanoparticles delivery approach provides:
1. Protection of the RNA cargo during development until it reaches the target cells.
2. Enhance uptake by antigen presenting cells.
3. A means to release the RNA cargo from the early endosomes by interacting with the endosomal membrane.
Figure 2. Non-viral saRNA delivery systems. Lipid-, polymer-, and emulsion-based delivery systems all use cationic groups to mediate condensation of the anionic RNA as well as delivery across the cell membrane. LNP systems, which have been found to be the most potent vaccine formulation, utilize a pH-sensitive ionizable cationic lipid as the main constituent and are taken up in cells through receptor-mediated endocytosis. In the endosome, the lower pH environment ionizes the cationic lipids, which then interacts electrostatically with anionic lipids in the endosomal membrane. These ion pairs cause a phase transition into a porous hexagonal phase (HII) that disrupts the endosome and facilitates release of the RNA into the cytoplasm.
Understanding the mechanism of LNP formation and delivery
Precision NanoSystems provides a range of technologies to accelerate the development of LNPs, including saRNA vectors, ionizable lipids and formulations, RUO reagent products, LNP manufacturing technologies, and biopharmaceutical services. One of Precision NanoSystems’s successful global academic partnerships highlighting the LNP technology expertise is a study conducted for Polymeric and lipid nanoparticles to deliver of self-amplifying RNA vaccines. The study highlights the success of this newer yet gold standard approach to vaccine development.
In the study conducted at Imperial College, London, researchers used saRNA encoding pre-fusion stabilized SARS-CoV-2 spike protein delivered as LNP for developing the COVID-19 vaccine and HA against influenza. Dr. Anna Blakney, the study's first author and now an Assistant Professor in the Michael Smith Laboratories and School of Biomedical Engineering at UBC, worked on the development of molecular and biomaterial engineering strategies for the delivery of self-amplifying RNA under the supervision of Prof. Robin Shattock and Prof. Molly Stevens of Imperial College, London. During this time, she and her research team aimed to better understand how the components of gene delivery formulations interact with the immune system to improve potency and enable clinical translation. They approached Dr. Andy Geal, Ex-CSO of Precision NanoSystems and now Chief Development Officer at Replicate Biosciences, and Dr. Anitha Thomas, Director of R&D at Precision NanoSystems, because they wanted to experiment with RNA formulation approaches, nanoparticle characterization methods, and with a variety of biochemical assays. Dr. Blakney developed the study using the NxGen™ microfluidic instrument for lipid nanoparticle formation-the Ignite™, and Dr. Anitha Thomas, who has extensive experience in LNP formulations, gave the research team access to PNI’s proprietary LNP library. The goal was to choose application-specific LNP compositions. Through their study, Dr. Blakney et al. provided an understanding of how saRNA formulated with LNP is a safe and efficient delivery vehicle for saRNA vaccines today.
In the study published in the Journal of Controlled Release, researchers extensively investigated the role of lipids and the route of administration (intramuscular versus intranasal). It was observed that both factors impacted the vaccine immunogenicity of two model antigens (influenza hemagglutinin and SARS-CoV-2 spike protein). They compared two delivery vehicles for saRNA vaccines, a lipid nanoparticle and a bio-reducible polymer (pABOL), specifically assessing protein expression and immune responses. For the LNP delivery system, they prepared a proprietary mix of ionizable lipids with two helper lipids (Precision NanoSystems, Inc.) at a total lipid concentration of 25 mM in ethanol. Precision NanoSystems research team assisted with selecting the right ionizable lipids and applied years of knowledge and expertise to formulate an application-specific mixture, narrowing the design space for a quicker, successful proof-of-concept generation.
It was observed that while the polyplex formulation had a higher protein expression, the LNP encapsulated saRNA exhibited higher humoral and cellular immunity in both vaccine models (influenza hemagglutinin and the SARS-CoV-2 spike glycoprotein). Although the size of the pABOL polyplexes and LNP were equivalent, the pABOL particles had a net positive surface charge. On the other hand, the LNP formulations had a neutral surface charge at physiological pH, as expected for ionizable cationic lipid comprising LNP, which is commonly the case for mRNA and saRNA LNP formulations. The researchers concluded that a neutral surface charge could have increased immunogenicity. It was also found that helper lipids also impact immunogenicity. While assessing the route of administration, it was observed that both systemic and mucosal antibodies developed in Intramuscular (IM) and Intranasal (IN) administrations; however, the responses were 10-fold higher using the IM route of administration. The findings suggested that different delivery systems and routes of administration may fill different delivery niches in the field of saRNA or mRNA-based vaccines. Overall, all groups of mice vaccinated (IM) with LNP formulations induced superior cellular immunity and reactogenicity. Although more research is needed in the pre-clinical phase, the findings suggest that the saRNA formulation with IM and IN route of administration can cause acute, cytokine-driven reactogenicity, allowing for potent humoral and cellular responses.
The study emphasizes the importance of LNP delivery vehicles and selecting appropriate formulations to ensure effective therapeutics or vaccines across applications. The successful use of Precision NanoSystems lipid formulations in this study demonstrates the company's years of expertise in LNP formulation and mRNA-LNPs production, which is easily scalable across its NanoAssemblr® family of instruments which operate on the same advanced NxGen™ microfluidics technology.
Figure 3. Development of LNP- mRNA Vaccine
LNPs, as a clinically validated and scalable technology, are the vaccine development technology of the future. To be prepared for pandemics, it is imperative that we conduct extensive research and investigate the potential of LNP delivered saRNA vaccines (Figure3), which are more efficient, require a lower dose, and are available for faster vaccine production.
Learn more about Precision NanoSystems’ COVID-19 saRNA-LNP vaccine in the virtual symposium Vaccines from Concept to Clinic: New insights into accelerating the development of RNA vaccines.
1. Blakney, A.K.; Ip, S.; Geall, A.J. An Update on Self-Amplifying mRNA Vaccine Development. Vaccines 2021, 9, 97.
2. Anna K. Blakney, Paul F. McKay, Kai Hu, Karnyart Samnuan, Nikita Jain, Andrew Brown, Anitha Thomas, Paul Rogers, Krunal Polra, Hadijatou Sallah, Jonathan Yeow, Yunqing Zhu, Molly M. Stevens, Andrew Geall, Robin J. Shattock. Polymeric and lipid nanoparticles for delivery of self-amplifying RNA vaccines. Journal of Controlled Release, Volume 338,2021, Pages 201-210.
Jun 09, 2022
Sep 28, 2022